JP2006105911A - Unidimensional measuring unit - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To easily position accurately a unidimensional measuring unit used for a measuring instrument using an evanescent wave. <P>SOLUTION: A guide groove 50e for conveyance extended linearly along a longitudinal direction is formed on a bottom face of a dielectric block 50 in the unidimensional measuring unit 10 having a large number of measuring flow passages 63 formed with a space along a longitudinal direction of the dielectric block 50, on a thin film layer of the slender dielectric block 50 formed with the thin film layer on a transparent and smooth upper face, and positioning abutting faces 50d, 50f are provided respectively on a side face and one end face to conduct the accurate positioning along X-, Y- and Z-directions. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention generates an evanescent wave by totally reflecting a light beam at an interface between a thin film layer in contact with a measurement object such as a sample and a dielectric block, thereby measuring a change in the intensity of the totally reflected light beam. In particular, the present invention relates to a measurement unit used in a measurement apparatus for analyzing a sample, and more particularly, to an elongated one-dimensional measurement unit including a large number of measurement units arranged in the length direction.

  Conventionally, a surface plasmon sensor is known as one of measuring devices using evanescent waves. In the metal, free electrons collectively vibrate to generate a dense wave called a plasma wave. A quantized version of this dense wave generated on the metal surface is called surface plasmon. The surface plasmon sensor analyzes the characteristics of a sample using a phenomenon in which the surface plasmon is excited by a light wave, and various types of sensors have been proposed. Among them, one that uses a system called Kretschmann configuration is well known (for example, see Patent Document 1).

  The surface plasmon sensor using the above system basically generates a light beam, for example, a dielectric block formed in a prism shape, a metal film formed on one surface of the dielectric block and brought into contact with a sample, and the like. A light source, an optical system for making the light beam incident at various angles so that a total reflection condition is obtained at the interface between the dielectric block and the metal film, and a light detection means for detecting the intensity of the light beam totally reflected at the interface And measuring means for measuring the surface plasmon resonance state based on the detection result of the light detecting means.

  In order to obtain various incident angles as described above, a relatively thin light beam may be incident on the interface by changing the incident angle, or a component incident on the light beam at various angles is included. As described above, a relatively thick light beam may be incident on the interface in a convergent light state or a divergent light state. In the former case, a light beam whose reflection angle changes according to the change in the incident angle of the incident light beam is detected by a small photodetector that moves in synchronization with the change in the reflection angle, or the direction in which the reflection angle changes Can be detected by an area sensor extending along the line. On the other hand, in the latter case, it can be detected by an area sensor extending in a direction in which each light beam reflected at various reflection angles can be received.

In the surface plasmon sensor having the above configuration, when a light beam is incident on the metal film at a specific incident angle θ _SP that is equal to or greater than the total reflection angle, an evanescent wave having an electric field distribution is generated in the sample in contact with the metal film. Surface plasmons are excited at the interface between the metal film and the sample by the evanescent wave. When the wave number vector of the evanescent light is equal to the wave number of the surface plasmon and the wave number matching is established, both are in a resonance state, and the light energy is transferred to the surface plasmon, so that the total reflection is made at the interface between the dielectric block and the metal film. The intensity of the light is sharply reduced. This decrease in light intensity is generally detected as a dark line by the light detection means.

When the wave number of the surface plasmon is found from a specific incident angle θ _SP (hereinafter referred to as total reflection attenuation angle θ _SP ) that is greater than or equal to the total reflection angle at which the light intensity is reduced, the dielectric constant of the sample is obtained. That is, when the surface plasmon wave number is K _SP , the surface plasmon angular frequency is ω, c is the speed of light in vacuum, εm and εs are metal, and the dielectric constant of the sample is as follows.

Knowing the dielectric constant εs of the sample, the refractive index or the like of the sample is found based on a predetermined calibration curve or the like, after all, by knowing the total reflection attenuation angle theta _SP, dielectric constant, that of the sample related to the refractive index Characteristics can be obtained.

  Further, a leak mode sensor is also known as a similar sensor using an evanescent wave (see, for example, Non-Patent Document 1). This leakage mode sensor basically includes, for example, a dielectric block formed in a prism shape, a clad layer formed on one surface of the dielectric block, and formed on the clad layer to be brought into contact with a sample. Optical waveguide layer, a light source that generates a light beam, an optical system that makes the light beam incident at various angles so that a total reflection condition is obtained at the interface between the dielectric block and the cladding layer, and total reflection at the interface A light detection means for measuring the intensity of the light beam and a measurement means for measuring the excited state of the waveguide mode based on the detection result of the light detection means are provided.

In the leakage mode sensor configured as described above, when a light beam is incident on the cladding layer through the dielectric block at an incident angle greater than the total reflection angle, the light waveguide layer transmits a specific wave number after passing through the cladding layer. Only light having a specific incident angle is propagated in the guided mode. When the waveguide mode is excited in this way, most of the incident light is taken into the optical waveguide layer, resulting in total reflection attenuation in which the intensity of light totally reflected at the interface is sharply reduced. Since the wave number of the waveguide light depends on the refractive index of the sample on the optical waveguide layer, by knowing the total reflection attenuation angle theta _SP, it can be analyzed refractive index of the sample and the properties of the sample related thereto .

In addition, the surface plasmon sensor and leakage mode sensor described above may be used for random screening to find a specific substance that binds to a desired sensing substance in the field of drug discovery research, etc. In this case, the thin film layer (surface In the case of a plasmon sensor, it is a metal film, and in the case of a leak mode sensor, a sensing substance is fixed on the clad layer and the optical waveguide layer), and various analyte solutions (liquid samples) are added on the sensing substance, every time a predetermined time elapses to measure the attenuated total reflection angle theta _SP described above. If the analyte in the liquid sample binds to the sensing substance, the refractive index of the sensing substance changes with time due to this binding. Therefore, the attenuated total reflection angle theta _SP was measured every predetermined time, by a change in the attenuated total reflection angle theta _SP to measure whether occurring, or is made binding between a test substance and a sensing substance whether That is, it can be determined whether or not the subject is a specific substance that binds to the sensing substance. Examples of such a combination of a specific substance and a sensing substance include an antigen and an antibody or an antibody and an antibody. As a specific measurement related to such a substance, for example, the sensing substance is a rabbit anti-human IgG antibody. In addition, there is a measurement for detecting the presence or absence of binding to a human IgG antibody as a specimen and quantitative analysis thereof.

In order to measure a binding state between a test substance and a sensing substance in the liquid sample, it is not always necessary to detect the angle itself of an attenuated total reflection angle theta _SP. For example, adding a liquid sample containing the analyte to the sensing substance, by measuring the angle variation subsequent ATR angle theta _SP, it is also possible to measure a binding state based on the magnitude of the angle variation .

  On the other hand, as a sensor as described above, a sensor that performs measurement by continuously supplying a liquid sample using a flow path mechanism on a measurement chip on a flat plate on which a sensing substance is fixed is known (for example, Patent Document 2). If a sensor of this form is used, when measuring the binding state between the sensing substance and the specific substance, a new liquid sample is always supplied onto the measurement chip, so the concentration of the analyte in the liquid sample does not change, The coupled state can be measured with high accuracy. In addition, after the binding state of the sensing substance and the specific substance is measured, if binding is performed, a liquid sample that does not contain the specific substance is allowed to flow over the measurement chip to which the conjugate is fixed. Thus, the dissociation state between the sensing substance and the specific substance can be measured. Furthermore, for example, when a gas is used as the sample, or when a liquid sample in which a gas is dissolved is used, the sample can be easily supplied onto the measurement chip using the flow path mechanism.

  Furthermore, in recent years, with the diversification of the reaction to be detected, various solvents have been used as measurement sample solvents, and some of these solvents, such as water, are relatively easy to evaporate. In this case, the evaporation of water means a change in the refractive index of the measurement sample, and the measurement signal also changes, so that accurate measurement may be difficult. By providing the flow path mechanism, it is possible to minimize the evaporation of the measurement sample and stabilize the signal.

Various effects can be obtained by providing the flow path mechanism in this way, but on the other hand, a long pipe is required to supply the sample on the measurement chip, and a large amount of sample must be prepared. There is also.

Therefore, the applicant of the present invention closely contacts the flow path forming member on the thin film layer of the elongated dielectric block in which the thin film layer is formed on the upper surface that is transparent and smooth with respect to the light beam. A number of flow paths are formed at intervals in the length direction of the dielectric block, the flow paths are used as measurement paths on the thin film layer, and both ends of each measurement path are connected to the inlet and outlet on the flow path forming member. A so-called one-dimensional measurement unit has been proposed in which a liquid sample is supplied and discharged by communicating with each other as a supply channel and a discharge channel with pipettes facing the inlet and the outlet, respectively. (Japanese Patent Application No. 2004-246879)
JP-A-6-167443 JP 2000-065731 A “Spectroscopy” Vol. 47, No. 1 (1998)

  Such a one-dimensional measurement unit can supply a liquid sample directly from an inlet of the flow path forming member by an external liquid feeding component such as a pipette tip, and thus has a flow path mechanism for supplying the sample onto the thin film layer. Thus, a long pipe or the like that has been conventionally required is not required, and measurement can be performed with a small number of samples.

  However, in order to accurately set the position of the long measurement unit at a predetermined position, since the protrusions provided at both ends of the unit are held and handled, the holding mechanism has three of X, Y, and Z. It is necessary to provide a function for accurately performing the operation in the direction, and the holding mechanism becomes complicated, and position fluctuation at the time of reading tends to occur, which is not preferable in practice.

  In view of the above circumstances, the present invention can accurately position the long measurement unit in the three directions X, Y, and Z at a predetermined position without a complicated holding mechanism. It is an object of the present invention to provide a one-dimensional measurement unit in which position fluctuation at the time of reading does not occur.

The one-dimensional measurement unit of the present invention is transparent to a light beam, and has an elongated dielectric block having a thin film layer formed on a smooth upper surface, and is in close contact with the thin film layer of the dielectric block. A flow path forming member that forms a large number of flow paths at intervals in the length direction of the dielectric block, and each of the multiple flow paths is formed on a thin film layer, a flow path formation In the one-dimensional measurement unit configured by a supply path from the inlet on the member to the measurement path and a discharge path from the measurement path to the outlet on the flow path forming member,
The dielectric block has a conveying guide groove extending linearly in the length direction on the bottom surface, and a positioning contact surface on each of the side surface and one end surface.

  The positioning contact surfaces provided on the side surface and one end surface of the dielectric block are preferably formed on the side surface near the bottom surface and the one end surface near the bottom surface, respectively.

  The measurement unit of the present invention may be configured as a measurement unit used for a so-called surface plasmon sensor, in which a thin film layer is made of a metal film, and measurement is performed using the effect of the surface plasmon resonance described above. The layer is composed of a clad layer formed on the one surface of the dielectric block and an optical waveguide layer formed on the clad layer, and measurement is performed by using the effect of waveguide mode excitation in the optical waveguide layer. You may comprise as a measurement unit used for a leak mode sensor.

  The dielectric block allows the light beam emitted from the light source of the measurement apparatus to enter the interface between the dielectric block and the thin film layer, and directs the light beam totally reflected at the interface toward the light detection means of the measurement apparatus. It may be in the form of a plate that does not include the prism to be emitted, or may be formed integrally with the prism.

  Further, the flow path forming member is preferably made of an elastic material, and in this case, it is more preferable that the inlet and / or the outlet is provided with a slit portion or a septum. Here, the slit portion or the septum is not necessarily located at the end of the inlet and / or the outlet, but may be in the vicinity of the inlet and / or the outlet.

  Further, it is preferable to further include a holding member that engages with the dielectric block and holds the flow path forming member on one surface of the dielectric block. In this case, the holding member includes an inlet of the flow path forming member and The holding plate portion is in close contact with the surface on which the outlet is formed, and the holding plate portion has a tapered insertion hole that narrows toward the flow path forming member at a position facing the inlet and the outlet of the flow path forming member. It is preferable to have it.

  Moreover, it is preferable to further include an evaporation preventing member that seals the inlet and / or the outlet and prevents evaporation of the sample. In this case, the evaporation preventing member may be made of an elastic material in which a slit is formed in a portion facing the inlet and / or the outlet. Further, the holding member and the evaporation preventing member may be formed integrally or may be bonded by an adhesive member.

  Note that, when measuring a measurement object such as a sample using the measurement unit of the present invention, that is, when obtaining refractive index information of the measurement object, for example, the refraction of the sample arranged on the thin film layer The rate itself may be acquired, or a sensing substance such as an antibody may be fixed on the thin film layer, and the refractive index change due to the antigen-antibody reaction or the presence or absence of the change may be acquired.

  The method for obtaining the refractive index information is to detect the reflected light at the interface between the dielectric block and the thin film layer at various incident angles and detect the total reflection attenuation angle or change in the angle. It is also possible to acquire the refractive index or the refractive index change by detecting, and DVNoort, K. johansen, C.-F. Mandenius, Porous Gold in Surface Plasmon Resonance Measurement, EUROSENSORS XIII, 1999 , pp.585-588, a light beam having a plurality of wavelengths is incident at an incident angle at which the total reflection condition is obtained at the interface, and the intensity of the light beam totally reflected at the interface for each wavelength. And the refractive index or refractive index change may be obtained by detecting the degree of total reflection attenuation for each wavelength. Furthermore, as described in PINikitin, ANGrigorenko, AA Beloglazov, MVValeiko, AISavchuk, OASavchuk, Surface Plasmon Resonance Interferometry for Micro-Array Biosensing, EUROSENSORS XIII, 1999, pp.235-238 Is incident at an angle of incidence that provides a total reflection condition at the interface, and a part of the light beam is split before the light beam enters the interface, and the split light beam is totally reflected at the interface. A change in refractive index may be acquired by causing interference with a reflected light beam and detecting a change in interference fringes of the light beam after the interference.

  That is, the refractive index information of the measurement object may be any information as long as it changes according to the refractive index of the measurement object, and the total reflection attenuation angle and total reflection that change according to the refractive index of the measurement object. Examples include a light beam wavelength that causes attenuation, a change in total reflection attenuation angle that changes in accordance with a change in the refractive index of the measurement object, a change in light beam wavelength that causes total reflection attenuation, or a change in interference fringes.

  In the one-dimensional measuring unit of the present invention, an elongated dielectric block in which a number of flow paths are formed on a thin film layer by a flow path forming member at intervals in the length direction extends linearly on the bottom surface in the length direction. Because it has a guide groove for use, it can be moved stably in the length direction by sliding the guide groove for conveyance on the rail on the device side so that the measurement path of each flow path faces a predetermined measurement section. And the dielectric block has positioning contact surfaces on the side surface and one end surface thereof, so that a positioning stopper that contacts the contact surface on the apparatus side, and the dielectric block is connected to the stopper and the rail. If a means for pressing toward is provided, the dielectric block can be accurately positioned in the three directions of X, Y, and Z, and stable and accurate measurement can be realized.

  Measurement can be performed.

  In addition, if the positioning contact surfaces provided on the side surface and one end surface of the dielectric block are formed on the side surface near the bottom surface and the one end surface near the bottom surface of the dielectric block, the positioning stopper can be provided integrally with the rail. The configuration for positioning on the apparatus side can be made extremely simple.

  Since the flow path forming member is made of an elastic material, the flow path forming member can be reliably brought into close contact with the thin film layer, so that liquid leakage of the liquid sample from the contact surface can be prevented. Further, in this case, since the liquid sample can be prevented from evaporating by further providing a slit or a septum at the inlet and / or the outlet, the change in the refractive index of the sample due to the evaporation of the liquid sample can be prevented. Therefore, the measurement signal can be stabilized.

  Further, by further including a holding member that engages with the dielectric block and holds the flow path forming member on one surface of the dielectric block, the dielectric block and the flow path forming member can be separated from each other during transportation. Therefore, the handleability can be improved. Further, in this case, the holding member includes a holding plate portion that is in close contact with the surface on which the inlet and outlet of the flow path forming member are formed, and the holding plate portion is located at a position facing the inlet and outlet of the flow path forming member. By providing a tapered insertion hole that narrows toward the flow path forming member, an external liquid feeding component such as a pipette or a syringe can be easily inserted into the inlet or outlet of the flow path forming member.

  In addition, by providing an evaporation preventing member that seals the inlet and / or outlet to prevent evaporation of the sample, it is possible to prevent a change in the refractive index of the sample due to evaporation of the liquid sample. Can be achieved. In this case, the evaporation preventing member can be made with a simple structure by forming the evaporation preventing member from an elastic material having a slit formed in a portion facing the inlet and / or the outlet.

  Further, when the holding member and the evaporation preventing member are integrally formed, the number of parts can be reduced, so that the productivity of the measurement unit can be improved.

  Further, when the holding member and the evaporation preventing member are bonded by the adhesive member, both can be formed of different materials.

  In addition, by providing the flow path forming member with a plurality of flow paths, it is possible to measure a plurality of samples in parallel with one measurement unit.

  Hereinafter, embodiments of the present invention will be described with reference to the drawings.

[Measurement unit]
1 is a perspective view of a measurement unit according to a first embodiment of the present invention, FIG. 2 is an exploded perspective view of the measurement unit, FIG. 3 is a top view of the measurement unit, and FIG. 4 is IV-IV in FIG. It is line sectional drawing.

  The measurement unit 10 is transparent to the light beam and is in close contact with the dielectric block 50 in which a metal film 55 as a thin film layer is formed on a smooth upper surface 50a, and the metal film 55 of the dielectric block 50. The flow path forming member 51 and the holding member 52 that engages with the dielectric block 50 and holds the flow path forming member 51 on the upper surface 50a of the dielectric block 50 are configured.

  The dielectric block 50 is made of, for example, a transparent resin, and has a trapezoidal body whose section perpendicular to the longitudinal direction is shorter at the lower base than at the upper base, and at both ends in the longitudinal direction of the main body. A holding portion 50b having a width smaller than that of the main body when viewed from the upper surface (or lower surface) direction is formed, and a light beam emitted from a light source of a measuring apparatus described later is applied to the dielectric block 50 and the metal film. A prism portion is formed integrally so as to be incident on the interface with 55 and to emit the light beam totally reflected on the interface toward the light detection means of the measuring apparatus. On both side surfaces in the longitudinal direction of the main body, there are an engaging convex portion 50c for engaging with an engaging hole 52c formed in a holding member 52, which will be described later, and a vertical convex portion 50d having a side surface near the bottom surface formed vertically. Sliding grooves 50e extending in parallel with the longitudinal direction are formed on the bottom surface. The surfaces of the vertical protrusions 50d formed on the left and right side surfaces are parallel to each other. In addition, a positioning contact surface 50f is provided on the end surface of the holding portion 50b on the side that becomes the tip when moved to the measurement unit.

  The flow path forming member 51 includes a flow path 60 composed of a supply path 62 extending from the inlet 61 to the measurement path 63 and a discharge path 64 extending from the measurement path 63 to the outlet 65 in the longitudinal direction of the flow path forming member 51. The plurality of flow paths 60 are arranged in a straight line.

  As shown in FIG. 4, at the lower part of the flow path forming member 51, the outlet of the supply path 62 and the inlet of the discharge path 64 are opened, and the surface of the metal film 55 located on the lower surface of the flow path forming member 51 A seal portion 51a surrounding the outlet of the supply passage 62 and the inlet of the discharge passage 64 is formed in a contact area, and the inner side of the seal portion 51a becomes the measurement passage 63. For this reason, when the flow path forming member 51 is brought into close contact with the metal film 55 of the dielectric block 50, the measurement path 63 in the seal portion 51a functions as a flow path. The seal portion 51a may be integrally formed with the upper portion of the flow path forming member 51, or may be formed of a material different from the upper portion and attached later, for example, An O-ring or the like attached to the lower portion of the flow path forming member 51 may be used.

  In a measuring device such as a surface plasmon sensor using the measuring unit of the present invention, it is assumed that a liquid sample containing a protein is used. However, if a protein in the liquid sample is fixed in the flow channel 60, the measurement is performed. Therefore, it is preferable that the material of the flow path forming member 51 does not have non-specific adsorptivity to proteins, and specifically, silicon, polypropylene, or the like may be used. In addition, since the flow path forming member 51 is made of such an elastic material, the flow path forming member 51 can be reliably brought into close contact with the metal film 55, so that the liquid sample leaks from the contact surface. Can be prevented.

  The holding member 52 is made of an elastic material such as polypropylene, and the cross section in the direction orthogonal to the longitudinal direction has a substantially square shape, and the inlet of the flow path forming member 51 on the upper plate (holding plate portion) of the holding member 52 A tapered pipette insertion hole 52a that narrows toward the flow path forming member 51 is formed at a position opposite to the outlet 61 and the outlet 65. A positioning boss 52b is formed on the outer side of the pipette insertion hole 52a.

  Further, an evaporation preventing member 54 is attached to the upper surface of the holding member 52 with a double-sided tape (adhesive member) 53. As shown in FIG. 2, a pipette insertion hole 53a is formed at a position facing the pipette insertion hole 52a of the double-sided tape 53, and a positioning hole 53b is formed at a position facing the boss 52b. Similarly, a slit 54a is formed at a position facing the pipette insertion hole 52a of the evaporation preventing member 54, and a positioning hole 54b is formed at a position facing the boss 52b. In a state where the hole 53b and the hole 54b of the evaporation preventing member 54 are inserted, the evaporation preventing member 54 is attached to the upper surface of the holding member 52, so that the slit 54a of the evaporation preventing member 54, the inlet 61 and the outlet of the flow path forming member 51 are provided. 65 is configured to face each other. The evaporation preventing member 54 needs to be made of a material having elasticity so that a pipette can be inserted from the slit 54a, and specifically silicon or polypropylene may be used. Note that the holding member 52 and the evaporation preventing member 54 may be formed integrally, and in addition to this, the flow path forming member 51 may be formed integrally.

  Furthermore, an engagement hole 52c for engaging with an engagement protrusion 50c formed in the dielectric block 50 is formed in the longitudinal side plate of the holding member 52, and the engagement hole 52c is engaged with the engagement protrusion 52c. In a state where the holding member 52 and the dielectric block 50 are engaged with each other by the portion 50c, the flow path forming member 51 is sandwiched between the holding member 52 and the dielectric block 50, and the flow path forming member 51 is The body block 50 is configured to be held on the upper surface 50a.

  As shown in FIG. 4, when the flow path forming member 51 is sandwiched between the holding member 52 and the dielectric block 50, the inlet 61 and the outlet 65 of the flow path forming member 51 are formed by the slit 54a of the evaporation preventing member 54. The liquid sample that is blocked from the outside air and injected into the flow path 60 is configured to prevent evaporation.

[Surface plasmon sensor]
Next, a surface plasmon sensor using the measurement unit 10 will be described. 5 is a plan view showing a schematic configuration of the surface plasmon sensor, FIG. 6 is a plan view of a measurement system of the surface plasmon sensor, FIG. 7 is a sectional view taken along line VII-VII in FIG. 6, and FIG. It is a side view of the measurement system. In FIG. 8, the holding member 52 (including the double-sided tape 53 and the evaporation preventing member 54) of the measurement unit 10 is omitted.

  As shown in FIG. 5, the surface plasmon sensor 1 has a surface that can simultaneously analyze a plurality of samples by allowing a light beam to be incident in parallel on a plurality of flow paths 60 provided in the measurement unit 10. It is a plasmon sensor, and is composed of a plurality of surface plasmon measurement systems 1A, 1B,. The configuration of each measurement system will be described by omitting the symbols A, B,.

  As shown in FIGS. 6 and 8, each measurement system includes a light source 14 (hereinafter referred to as a laser light source 14) composed of a semiconductor laser or the like that generates a single light beam 13, and the light beam 13 is supplied to the measurement unit 10. Through the optical system 15 that is incident in parallel to the two interfaces 50d and 50e between the dielectric block 50 and the metal film 55 below the flow path 60 so as to obtain various incident angles, and the interface Two collimator lenses 16 that collimate the light beams 13 totally reflected by 50d and 50e, respectively, two photodiode arrays 17 that respectively detect the collimated light beams 13, and two photodiode arrays 17 A differential amplifier array 18, a driver 19, a signal processing unit 20 including a computer system, and a display unit 21 connected to the signal processing unit 20.

  The incident optical system 15 includes a collimator lens 15a that collimates the light beam 13 emitted from the laser light source 14 in a divergent light state, a half mirror 15c that separates the collimated light beam 13, and a half mirror 15c. A mirror 15d for reflecting the reflected light beam 13 in the direction of the measurement unit 10, a light beam 13 transmitted through the half mirror 15c, and a light beam 13 reflected by the mirror 15d are converged on the interfaces 50d and 50e, respectively. And two condenser lenses 15b.

  Since the light beam 13 is condensed as described above, the light beam 13 includes components incident on the interfaces 50d and 50e at various incident angles θ. In addition, this incident angle (theta) shall be an angle more than a total reflection angle. Therefore, the light beam 13 is totally reflected at the interfaces 50d and 50e, and the reflected light beam 13 includes components reflected at various reflection angles. The optical system 15 may be configured to allow the light beam 13 to enter the interfaces 50d and 50e in a defocused state. By doing so, errors in surface plasmon resonance state detection are averaged, and measurement accuracy is improved.

  The light beam 13 is incident on the interfaces 50d and 50e as p-polarized light. In order to do so, the laser light source 14 may be disposed in advance so that the polarization direction thereof is a predetermined direction. In addition, the direction of polarization of the light beam 13 may be controlled with a wave plate.

  As shown in FIG. 7, in the present embodiment, the light beam 13 is incident on the measurement paths 63 of the respective flow paths 60 of the measurement unit 10 in parallel with respect to the two interfaces 55d and 55e. Nothing is fixed on the metal film 55 on the inner interface 55d, and the sensing substance 73 is fixed on the metal film 55 on the interface 55e.

  In measurement, the differential value output by the differential amplifier of the detector of the measurement unit is continuously measured over time, and the change in the refractive index of the liquid sample 72 or sensing substance 73 in contact with the metal film 55 is examined. Can do.

  In particular, in this embodiment, if the analyte contained in the liquid sample is a specific substance that binds to the sensing substance 73, the refractive index of the sensing substance 73 changes according to the binding state between the sensing substance 73 and the analyte. By continuing to measure the differential value, it is possible to detect whether or not the analyte is a specific substance that binds to the sensing substance 73.

  Further, in the present embodiment, there is a region where the sensing substance 73 is not fixed on the metal film 55 and a region where the sensing substance 73 is fixed, and the reference measurement and the binding between the sensing substance 73 and the subject are performed. Since the measurement of the state is performed at the same time, by obtaining the difference between the measurement values in the two regions, it is possible to obtain a measurement result that offsets the measurement error caused by the influence of the temperature change of the liquid sample.

  Note that, as described above, the method of use of this embodiment is not limited to the measurement of the reference and the measurement of the binding state between the sensing substance 73 and the analyte. In addition, an aspect in which a measurement surface formed by another flow path is used for reference measurement, or an aspect in which reference measurement is not performed is possible.

[Measurement unit positioning]
Hereinafter, positioning of the measurement unit in the surface plasmon sensor 1 having the above configuration will be described. Prior to the measurement, the measurement unit 10 is moved from the temperature-controlled room 2 toward the measurement position on the chip holder 11. (A movement mechanism will be described later) A rail 11a that engages with a sliding groove 50e formed in the dielectric block 50 is formed in the chip holding portion 11, and high positional accuracy is obtained when the measurement unit 10 is moved. It can be secured. Further, after the measurement unit 10 is placed on the chip holding part 11, the dielectric block 50 is placed on the stopper 11b integrally standing on the right side of the chip holding part 11 in the figure. The vertical convex portion 50d formed on the opposite side of the dielectric block 50 is pressed in the X direction by an urging mechanism (not shown) so that the vertical convex portion 50d contacts with the measurement position on the chip holding portion 11. It is fixed and positioning is performed accurately in the X direction. At this time, there is a slight play in the positioning by the engagement between the sliding groove 50e and the rail 11a so that precise positioning in the X direction can be performed. Further, a flat plate-like or frame-like pressing member 12 is placed on the upper surface of the flow path forming member 51, and the measuring unit 10 is pressed in the Z direction from above to accurately measure the measuring unit 10 on the chip holding portion 11 in the Z direction. Position to. Further, as shown in FIG. 9, the tip end (right end in the figure) of the chip holder 11 abuts against the tip surface 50f of the holder 50b of the dielectric block 50 to position the measurement unit 10 in its moving direction, that is, in the Y direction. A stopper 11c is provided, and the measurement unit 10 is positioned in the Y direction by pressing the end face of the holding portion 50b of the dielectric block 50 at the rear end. In FIG. 9, the stopper 11b that contacts the vertical convex portion 50d of the dielectric block 50 is not shown. (Figures 16 and 17 show other examples)
[Sample analysis with surface plasmon sensor]
Thereafter, as shown in FIG. 7, a pipette tip 70 for supplying the liquid sample is inserted into the inlet 61 of the flow path forming member 51, and a pipette tip 71 for sucking the liquid sample is inserted into the outlet 65. After supplying the liquid sample 72 to the measurement path 63 of the flow path 60, the measurement is started.

  As shown in FIG. 8, the light beam 13 emitted from the laser light source 14 in a divergent light state is caused on the interfaces 50 d and 50 e between the dielectric block 50 and the metal film 55 below the measurement path 63 by the action of the optical system 15. Converge. At this time, the light beam 13 includes components incident on the interfaces 50d and 50e at various incident angles θ. In addition, this incident angle (theta) shall be an angle more than a total reflection angle. Therefore, the light beam 13 is totally reflected at the interfaces 50d and 50e, and the reflected light beam 13 includes components reflected at various reflection angles.

  After total reflection at the interfaces 50d and 50e, the two light beams 13 respectively collimated by the two collimator lenses 16 are detected by the two photodiode arrays 17, respectively. In the photodiode array 17 in this example, a plurality of photodiodes 17a, 17b, 17c,... Are arranged in a line, and in the traveling direction of the collimated light beam 13 in the plane shown in FIG. On the other hand, the photodiodes are arranged in a direction in which the parallel arrangement direction of the photodiodes is substantially perpendicular. Therefore, different photodiodes 17a, 17b, 17c,... Receive each component of the light beam 13 totally reflected at various reflection angles at the interfaces 50d and 50e.

  In the present embodiment, there is a region where the sensing substance 73 is not fixed on the metal film 55 and a region where the sensing substance 73 is fixed, and the reference measurement and the binding state between the sensing substance 73 and the analyte are Since the measurement is performed at the same time, by obtaining the difference between the measured values in the two regions, it is possible to obtain a measurement result that offsets the measurement error caused by the influence of the temperature change of the liquid sample.

  Here, the metal film 55 is illustrated as the reference measurement surface, but it is preferable that the metal film 55 has a function of not being combined with the substance to be measured in the liquid sample 72. As such a measurement surface, for example, alkylthiol, amino alcohol, amino ether or the like may be fixed to the reference measurement surface, and an antibody may be fixed to the measurement surface of the subject as a sensing substance.

  Note that, as described above, the method of use of this embodiment is not limited to the measurement of the reference and the measurement of the binding state between the sensing substance 73 and the analyte. In addition, an aspect in which a measurement surface formed by another flow path is used for reference measurement, or an aspect in which reference measurement is not performed is possible.

  In addition, the measuring device is not limited to a mode in which a plurality of surface plasmon measurement systems simultaneously measure all the channels provided in the measurement unit, and includes only one surface plasmon measurement system, It is good also as an aspect which measures the several flow path provided in the measurement unit sequentially by moving the position of a measurement unit relatively with respect to a measurement system.

[Overall configuration of surface plasmon sensor device]
Hereinafter, an outline of an embodiment of a surface plasmonsen that analyzes a sample using the one-dimensional measurement unit of the present invention, including an overall basic configuration including a transport mechanism of the measurement unit, and a transport mechanism of the measurement unit An example will be described.

  10 is a plan view showing an outline of the entire surface plasmon sensor device, FIG. 11 is a front view thereof, FIG. 12 is a side view thereof, FIG. 13 is a perspective view showing a transport mechanism of a measurement unit, and FIG. 14 is a plan view thereof. FIG. 15 is a partial front view showing the measuring unit push-up mechanism.

  In the surface plasmon sensor device according to this embodiment, two unit plates 100 in which eight one-dimensional measuring units 10 (hereinafter referred to as units) are accommodated in parallel are arranged in parallel, and 16 units 10 can be measured at a time. It is configured as follows. The plate 100 is movable in the A direction by a distance of 16 in a direction perpendicular to the length direction of each unit 10, and all 16 units 10 are perpendicular to the A direction at the push-up position a described later. By being moved in the B direction, all the measurement paths of the unit 10 can be moved to the measurement unit 111 of the unit. Unlike the pressing member 12 in FIG. 9 that holds the unit 10 over the entire length of the unit 10, the measuring unit 111 has a well holding unit 112 that presses only the portion of the unit 10 located in the measuring unit 111 in the Z direction. Position the measurement path to be measured.

  A dispensing head 120 having tips 121 and 122 for supplying and discharging (inhaling) liquid can be moved in and out of the measuring unit 111. The dispensing head 120 is arranged from the top in FIG. It is possible to move in the A direction to the bottom. At the top of the figure, there is a chip supply unit 122 in which a large number of chips 121 are arranged vertically and horizontally. This chip supply unit 122 moves in the B direction and all the chips 121 move in the A direction. It can be picked up by the head 120. In addition, a titer plate 123 in which a large number of analytes used in measurement alongside the tip supply unit 122 are arranged vertically and horizontally is also installed so as to be movable in the B direction, and the trajectory in which the dispensing head 120 moves in the A direction. Moving in the direction B, all the analytes are inhaled by the tip 121 of the dispensing head 120. As this titer plate, for example, a 386 titer plate that dispenses 96 rows every other row and column can be used.

  Further, in the lower part of FIG. 10, DMSO solutions having different concentrations, for example, low concentration, medium low concentration, medium concentration, medium high concentration, high concentration DMSO solution, and 96 well plate 124 containing the buffer solution are similarly provided in the B direction. The sample is moved in the direction B, and all the samples and the buffer solution are sucked by the tip 121 of the dispensing head 120.

  In addition, between the 96 well plate 124 and the measuring unit 111, a waste liquid part 125 is disposed, which sucks out the unnecessary discarded liquid discharged from the unit from the pipette tip and discards the liquid. The head 120 discards the liquid here.

  The measurement order is as follows. First, the unit 10 in which a sensing substance is previously provided on a part of the metal film 55 is moved from the unit plate 100 to the measurement unit 111 in the B direction, and the measurement path 63 to be measured is positioned in the measurement unit 111. After fixing, a new tip 121 is attached to the dispensing head 120 from the tip supply unit 122, the head 120 moves to the 96-well plate in the direction A and sucks the low-concentration DMSO solution into the measuring path 63 of the unit 10. This low concentration DMSO solution is injected, measured, and then discharged. Next, similarly, inhalation, injection, measurement, and discharge are performed for the DMSO solution of medium concentration and high concentration. In this way, the DMSO concentration is changed and measurement for DMSO correction is performed. Thereafter, the head 120 moves to the analyte titer plate 123 to suck in the analyte, moves in the direction A, injects it into the unit 10, waits for the reaction, performs measurement, and discharges. Thereafter, the head 120 moves to the 96-well plate 124 to suck in the buffer solution, inject it into the unit 10, measure it, and discharge it. Then, it is washed with a buffer and the chip 121 is discarded. From the obtained DMSO concentration data, the refractive index of the analyte is obtained.

  This completes the measurement of one analyte for one measurement path with a sensing substance, and then moves the unit 10 in the B direction to position the next measurement path 63 in the measurement unit 111 and perform the same measurement. For different analytes.

  As shown in FIG. 11, two pairs of chips 121 are paired. This is because a pair of chips 121 enters the inlet 61 and the outlet 65 of the unit 10 at the same time to supply liquid from the inlet 61. This is for discharging the liquid from the outlet 65 at the same time.

  Also, in FIG. 12, above the chip supply unit 122 and the analyte titer plate 123 and above the 96-well plate 124, the chip 121 moves up and down in the apparatus, and each of the A chip opening 126 and a sample opening 127.

  Also, below the measurement unit, a laser light source 130, an optical system 132 including a beam splitter and a shutter for allowing the laser light to enter the unit 10, an optical fiber 131 connecting between them, and light reflected by the metal film 55 of the unit 10 are reflected. A light receiving unit 133 for detection is installed.

[Measurement unit transport]
A mechanism for moving the unit 10 is shown in FIGS. Two unit plates 100 accommodating eight one-dimensional measuring units 10 each (empty in the figure) can be moved in the A direction, and the unit 10 pushed upward by a push-up mechanism from the two unit plates 100 can be moved in the B direction. The measurement path to be measured is positioned in the measurement unit 111. As shown in FIG. 15, the push-up mechanism includes a push-up plate 140 that pushes up the unit 10 from the bottom to the top, and this push-up plate 140 is placed under the plate 100 of the unit and at the bottom of the plate 100 for each unit. The unit 10 is provided so as to push upward from the opened slit 100a (FIGS. 13 and 14). A pinion 143 meshes with a rack 142 fixed integrally with the lower portion 141 of the push-up plate 140, and a worm wheel 144 coaxial with the rack 142 is rotated by the worm gear 145. It is supposed to be.

  Details of the positioning mechanism of the unit 10 in this embodiment of the apparatus are shown in FIGS. A side stopper 40 abuts on the vertical convex portion 50d on the side surface of the unit 10, and a tip stopper 41 abuts on the tip surface 50f of the holding portion 50b. As shown in FIG. The side pressing portion 42 that contacts and positions the vertical convex portion 50d on the opposite side pushes the dielectric block 50 of the unit 10 from the opposite side at a position where the measurement path comes to the measuring portion 111, thereby blocking the block 50 on the side stopper. Press 40 and position unit 10 accurately.

  According to the positioning mechanism of the above apparatus, it is possible to perform accurate positioning, and the unit 10 can be fixed with a strong force. Therefore, there is an effect that the unit 10 does not move when the chip 121 is inserted and removed. .

The perspective view of the measurement unit by the 1st Embodiment of this invention Exploded perspective view of the measurement unit Top view of the above measurement unit IV-IV sectional view in FIG. The top view which shows schematic structure of the surface plasmon sensor using the measurement unit by the 1st Embodiment of this invention Plan view of the measurement system of the above surface plasmon sensor VII-VII line sectional view in FIG. Side view of the measurement system of the above surface plasmon sensor Front view showing the positioning direction of the measurement unit in the measurement system of the surface plasmon sensor The top view which shows the outline of the whole surface plasmon sensor apparatus Front view showing the outline of the entire surface plasmon sensor device Side view showing the outline of the entire surface plasmon sensor device The perspective view which shows the conveyance mechanism of a measurement unit Plan view showing transport mechanism of measurement unit Partial front view showing the measurement unit push-up mechanism of the transport mechanism of the measurement unit Partially enlarged perspective view showing the positioning mechanism of the measurement unit Partially enlarged side view showing the positioning mechanism of the measurement unit

Explanation of symbols

10 Measuring unit
12 Press member
13 Light beam
14 Laser light source
15 Optical system
16 Collimator lens
17 Photodiode array
17a, 17b, 17c …… Photodiode
18 Differential amplifier array
18a, 18b, 18c ... Differential amplifier
19 Drivers
20 Signal processor
21 Display
40 Side stopper
41 Tip stopper
42 Side pressing part
50 dielectric block
50d Vertical protrusion (side contact surface)
50f Tip contact surface
51 Channel formation member
52 Holding member
53 Double-sided tape
54 Evaporation prevention member
55 Metal film
60 channels
61 entrance
62 Supply channel
63 Measuring path
64 Discharge channel
65 Exit
70, 71 pipettes
72 Liquid sample
73 Sensing substances
100 unit plate
111 Measurement unit
120 dosing head
121 Tip (pipette)
122 Chip supply unit
123 Analyte titer plate
124 96 well plate (sample plate)
125 Liquid absorber
140 Push-up plate

Claims (2)

An elongated dielectric block transparent to the light beam and having a thin film layer formed on a smooth upper surface;
It is in close contact with the thin film layer of the dielectric block, and comprises a flow path forming member that forms a large number of flow paths on the thin film layer at intervals in the length direction of the dielectric block,
Each of the plurality of flow paths has a measurement path formed on the thin film layer, a supply path from the inlet on the flow path forming member to the measurement path, and from the measurement path to an outlet on the flow path forming member. It consists of a discharge path that leads to
The one-dimensional measuring unit, wherein the dielectric block has a conveyance guide groove extending linearly in the length direction on the bottom surface, and a positioning contact surface on each of the side surface and one end surface. .   The one-dimensional structure according to claim 1, wherein positioning contact surfaces provided on a side surface and one end surface of the dielectric block are formed on a side surface near the bottom surface and one end surface near the bottom surface, respectively. Measuring unit.

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